Persea americana Mill. (avocado fruit) has many health benefits when added to our diet due to various pharmacological activities, such as preventing bone loss and inflammation, modulating immune response and acting as an antioxidant. In the current study, the total ethanol extract (TEE) of the fruit was investigated for in vitro antioxidant and anti-inflammatory activity via DPPH and cyclooxygenase enzyme inhibition. Biological evaluation of the antiarthritic effect of the fruit extract was further investigated in vivo using Complete Freund's Adjuvant (CFA) arthritis model, where the average percentages of body weight change, inhibition of paw edema, basal paw diameter/weight and spleen index were estimated for all animal groups. Inflammatory mediators such as serum IL-6 and TNF-α were also determined, in addition to histopathological examination of the dissected limbs isolated from all experimental animals. Eighty-one metabolites belonging to different chemical classes were detected in the TEE of P. americana fruit via UPLC/HR-ESI-MS/MS. Two classes of lyso-glycerophospholipids; lyso-glycerophosphoethanolamines and lysoglycerophosphocholines were detected for the first time in avocado fruit in the positive mode. The TEE of fruit exhibited significant antioxidant and anti-inflammatory activity in vitro. In vivo anti-arthritic activity of the fruit TEE improved paw parameters, inflammatory mediators and spleen index. Histopathological findings showed marked improvements in the arthritic condition of the excised limbs. Therefore, avocado fruit could be proposed to be a powerful antioxidant and antiarthritic natural product.